Project Summary Preterm survivors acquire a variety of chronic hypoxia-ischemia (H-I)-related motor and neurobehavioral disabilities that have been widely assumed to arise from destructive cerebral gray and white matter lesions that deplete neurons and glia. Our recent human and experimental findings support an alternative primary mechanism whereby diffuse disturbances in cerebral development arise from disrupted maturation of pre- oligodendrocytes (preOLs) in white matter tracts and immature neurons in multiple cerebral regions. We will employ a novel instrumented preterm fetal sheep model of cerebral H-I that reproduces many key features of acute and chronic cerebral injury in human preterm survivors. With this model, we demonstrated that chronic white matter injury (WMI) results in myelination failure that arises from maturation arrest of preOLs that fail to differentiate and myelinate. Chronic WMI is accompanied by progressive volume loss of cortical and subcortical gray matter structures that arises from reduction in the basal dendritic arbor of immature projection neurons. We will test the over-riding hypothesis that cerebral H-I causes widespread disturbances in brain development through: (1) hypo-myelination of white matter tracts that integrate neuronal networks and (2) disturbances in the structural and functional maturation of neurons in multiple structures critical for learning, memory and cognition. To define the mechanisms of impaired white matter growth and myelination failure in chronic WMI, we will promote maturation of preOLs in vivo by blocking the hyaluronidase, PH20 with a novel inhibitor, SuBr3 (aim 1). We recently demonstrated in vitro that SuBr3 promotes preOL maturation and blocks the generation of digestion products of hyaluronic acid (HA) derived from PH20. We will also undertake mechanistic studies with SuBr3 to test the hypothesis that a reduction in myelination failure will promote maturation of the dendritic arbor of cortical projection neurons. We will thereby determine if there is a mechanistic link between myelination failure and cortical neuronal dysmaturation. Our preliminary studies support that the spectrum of neuronal dysmaturation varies markedly between the cortex and hippocampus. Whereas cortical pyramidal cells display reduced synaptic activity, hippocampal projection neurons display enhanced synaptic activity. Hence, defining the response to future regeneration and repair strategies will require a region-dependent analysis. Thus, we will undertake an integrated morphometric and physiological analysis at the single cell level of dysmature projection neurons in the hippocampus (aim 2) and cortex (aim 3). Our long-term objective is to develop therapies that promote myelination in chronic WMI without adverse effects on neuronal maturation. The proposed studies will provide rigorous outcome measures to quantify the response to a wide range of clinically feasible future potential therapies (e.g., optimized preterm nutrition or reduced neonatal stress) that have the potential to promote cerebral growth and reduce the broad and complex spectrum of life-long motor and neurobehavioral disabilities that are now common in human preterm survivors.